3-(D-gluconoylamino)propyl(2-hydroxyethyl)dimethylammonium chloride

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experimental starting date :October 4, 2017 - Experimental completion date : October 6, 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Version / remarks:

July 22, 2010

Deviations:

no

Remarks:

Only minor modifications.

GLP compliance:

no

Remarks:

Laboratory works with standard operating procedures and according to GLP procedures with the exception of audits by a separate Quality Assurance Unit.

Test material

Test material information

Constituent 1

Reference substance name:

3-(D-gluconoylamino)propyl(2-hydroxyethyl)dimethylammonium chloride

EC Number:

257-440-3

EC Name:

3-(D-gluconoylamino)propyl(2-hydroxyethyl)dimethylammonium chloride

Cas Number:

51812-80-7

Molecular formula:

C13H29N2O7.Cl

IUPAC Name:

1-Propanaminium, 3-(D-gluconoylamino)-N-(2-hydroxyethyl)-N,N-dimethyl-, chloride

Test material form:

liquid

Details on test material:

Batch 0002042127

Specific details on test material used for the study:

Solid fraction: 60.2%
Physical state: Clear yellow to light amber liquid
pH: 4.9
Storage at room temperature

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

foreskin from a single donor

Justification for test system used:

Certified by Ashland Tissue Engineering laboratories, October 2, 2017.

Vehicle:

unchanged (no vehicle)

Remarks:

Product is 60% solids in water

Details on test system:

- Ashland 0.5 cm2 reconstructed epidermis from normal human keratinocytes.
- Cells are grown on inert polycarbonate filter on chemically defined medium, airlifted for 17 days
- 3 tissues will be used for each chemical.
- The RHE tissues are incubated at 37°C, 5% CO2 until test substance application (usually 24 hours).

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

Undiluted

Duration of treatment / exposure:

42 minutes
Procedure:
. Fill a 24-well plate with 300μl by well of pre-warmed maintenance culture medium.
· Transfer tissues.
· Dispense 16 μL ± 0.5 μL (or 16 μg) of products and controls on the top of epidermis.
· Carefully apply a nylon mesh (Ø = 7.5mm) on the whole surface with forceps.
· Keep the plates containing the treated epidermis for 42 minutes at room temperature.

Duration of post-treatment incubation (if applicable):

42 hours and MTT incubation for 3 hours.
Procedure:
· Remove the nylon mesh with fine forceps from the epidermal surface of a treated tissue.
· Remove the treated units using forceps, and rinse thoroughly 25 times with 1 ml DPBS.
· Transfer the washed tissue on 2 mL growth culture medium (6-well plate).
· Incubate the treated and rinsed epidermis at 37°C, 5% CO2 for 42 hours (± 60 minutes).
Assessment of non-specific MTT reduction (NSMTT)
· To identify direct MTT reducers, each test chemical should be added to freshly prepared MTT solution. If the MTT mixture containing the test chemical turns blue/purple, the test chemical is presumed to directly reduce MTT and a further functional check on non-viable RhE tissues should be performed.
· Each MTT reducing test chemical is applied on at least two killed tissue replicates which undergo the entire testing procedure to generate a non-specific MTT reduction (NSMTT)

Number of replicates:

24-well plates

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Main Ashland Equivalent Epidermis Skin Irritation Test

Value:

91.12

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Remarks:

viability: 100%

Positive controls validity:

valid

Remarks:

viability: 3.08%

Remarks on result:

no indication of irritation

Any other information on results incl. tables

Cell viability results :

Group	Mean Viability (%)	Classification
Negative Control / Water	100	Non irritant
Positive Control / SDS 5%	3.08	Irritant
Tested Product / Neat CERAPHYL™ 60	91.12	Non irritant

 

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

According to the in-vitro Ashland Epidermis skin irritation test, CERAPHYL® 60 is classified as non-irritant.